This invention relates to inhibitors of cysteine proteases, in particular to dipeptide nitrile cathepsin K inhibitors and to their pharmaceutical use for the treatment or prophylaxis of diseases or medical conditions in which cathepsin K is implicated.
Cathepsin K is a member of the family of lysosomal cysteine cathepsin enzymes, e.g. cathepsins B, K, L and S, which are implicated in various disorders including inflammation, rheumatoid arthritis, osteoarthritis, osteoporosis, tumors (especially tumor invasion and tumor metastasis), coronary disease, atherosclerosis (including atherosclerotic plaque rupture and destabilization), autoimmune diseases, respiratory diseases, infectious diseases and immunologically mediated diseases (including transplant rejection).
Our copending International patent application WO 99/24460 describes dipeptide nitrites which are inhibitors of cysteine cathepsins and their use for treatment of cysteine cathepsin dependent diseases or medical conditions. New dipeptide nitrile compounds have now been made which are inhibitors of cathepsin K, and which have desirable properties for pharmaceutical applications.
Accordingly the present invention provides a compound of formula I, or a pharmaceutically acceptable salt or ester thereof: 
In which
R1 and R2 are independently H or C1-C7lower alkyl, or R1 and R2 together with the carbon atom to which they are attached form a C3-C8cycloalkyl ring, and
Het is an optionally substituted nitrogen-containing heterocyclic substituent, provided that Het is not 4-pyrrol-1-yl.
The Het substituent may be at the 2- or 3-position of the phenyl ring, though is preferably at the 4-position.
In the present description xe2x80x9cnitrogen-containing heterocyclexe2x80x9d signifies a heterocyclic ring system containing at least one nitrogen atom, from 2 to 10, preferably from 3 to 7, most preferably 4 or 5, carbon atoms and optionally one or more additional heteroatoms selected from O, S or preferably N.
Het may comprise an unsaturated, e.g. an aromatic, nitrogen-containing heterocycle; though preferably comprises a saturated nitrogen-containing heterocycle. Particularly preferred saturated nitrogen-containing heterocycles are piperazinyl, preferably piperazin-1-yl, or piperidinyl, preferably piperidin-4-yl.
Het may be substituted by one or more substituents, e.g. by up to 5 substituents independently selected from halogen, hydroxy, amino, nitro, optionally substituted C1-4alkyl (e.g. alkyl substituted by hydroxy, alkyloxy, amino, optionally substituted alkylamino, optionally substituted dialkylamino, aryl or heterocyclyl), C1-4alkoxy.
Preferably Het is substituted at a nitrogen atom, most preferably mono-substituted at a nitrogen atom.
Preferred substituents for Het are C1-C7lower alkyl, C1-C7lower alkoxy-C1-C7lower alkyl, C5-C10aryl-C1-C7lower alkyl, or C3-C8cycloalkyl.
R1 and R2 as C1-C7lower alkyl are preferably the same, e.g. methyl, or R1 and R2 together with the carbon atom to which they are attached preferably form a C3-C8cycloalkyl ring, e.g. a cyclopropyl ring. Most preferably both R1 and R2 are H.
Thus in particularly preferred embodiments the invention provides a compound of formula II, or a pharmaceutically acceptable salt or ester thereof: 
wherein X is CH or N, and
R is H, C1-C7lower alkyl, C1-C7lower alkoxy-C1-C7lower alkyl, C5-C10aryl-C1-C7lower alkyl, or C3-C8cycloalkyl.
Thus particular examples of R as C1-C7lower alkyl are methyl, ethyl, n-propyl, or i-propyl.
A particular example of R as C1-C7lower alkoxy-C1-C7lower alkyl is methoxyethyl.
A particular example of R as C5-C10aryl-C1-C7lower alkyl is benzyl.
A particular example of R as C3-C8cycloalkyl is cyclopentyl.
Examples of particular compounds of formula II are:
N-[1-(Cyanomethyl-carbamoyl)-cyclohexyl]-4-(piperazin-1-yl)-benzamide;
N-[1-(Cyanomethyl-carbamoyl)-cyclohexyl]-4-(4-methyl-piperazin-1-yl)-benzamide;
N-[1-(Cyanomethyl-carbamoyl)-cyclohexyl]-4-(4-ethyl-piperazin-1-yl)-benzamide;
N-[1-(Cyanomethyl-carbamoyl)-cyclohexyl]-4-[4-(1-propyl)-piperazin-1-yl]-benzamide;
N-[1-(Cyanomethyl-carbamoyl)-cyclohexyl]-4-(4-isopropyl-piperazin-1-yl)-benzamide;
N-[1-(Cyanomethyl-carbamoyl)-cyclohexyl]-4-(4-benzyl-piperazin-1-yl)-benzamide;
N-[1-(Cyanomethyl-carbamoyl)-cyclohexyl]-4-[4-(2-methoxy-ethyl)-piperazin-1-yl]-benzamide;
N-[1-(Cyanomethyl-carbamoyl)-cyclohexyl]-4-(1-propyl-piperidin-4-yl)-benzamide;
N-[1-(Cyanomethyl-carbamoyl)-cyclohexyl]-4-[1-(2-methoxy-ethyl)-piperidin-4-yl]-benzamide;
N-[1-(Cyanomethyl-carbamoyl)-cyclohexyl](1-isopropyl-piperidin-4-yl)-benzamide;
N-[1-(Cyanomethyl-carbamoyl)-cyclohexyl]-4-(1-cyclopentyl-piperidin-4-yl)-benzamide;
N-[1-(Cyanomethyl-carbamoyl)-cyclohexyl]-4-(1-methyl-piperidin-4-yl)-benzamide, and
N-[1-(Cyanomethyl-carbamoyl)-cyclohexyl]-4-(piperidin-4-yl)-benzamide.
Compounds of formula I and II and the specific compounds above are hereinafter referred to as Compounds of the Invention.
Compounds of the Invention may be prepared by coupling the corresponding Het substituted benzoic acid derivative with I-amino-cyclohexanecarboxylic acid cyanomethyl amide. For example, the benzoic acid derivative, preferably in the form of its hydrochloride, is mixed with 1-amino-cyclohexanecarboxylic acid cyanomethyl amide, e.g. in the presence of HOBT (1-hydroxybenzotriazole), WSCD and trimethylamine, in solution, e.g. in DMF, and stirred, e.g. overnight at room temperature. The product may be recovered, for instance, by evaporation of the solvent, followed by washing with aqueous sodium carbonate solution, preferably under mildly basic conditions, followed by solvent extraction, e.g. with ethyl acetate, drying of the extract, e.g. over sodium sulfate, evaporation of the solvent and filtration. Alternative procedures and reagents may be used; for instance, as hereinafter described in the Examples.
Thus in a further aspect the invention provides a process for the preparation of a compound of formula I which comprises coupling the corresponding Het substituted benzoic acid derivative of formula III: 
With 1-amino-cyclohexanecarboxylic acid cyanomethyl-amide. 
1-Amino-cyclohexanecarboxylic acid cyanomethyl-amide may be prepared by coupling 1-amino-cyclohexane carboxylic acid, typically in appropriate amino protected form, e.g. FMOC-1-amino-cyclohexane carboxylic acid, with 2-aminoacetonitrile. For example, FMOC-1-amino-cyclohexane carboxylic acid, e.g. with HOBT and WSCD, is added to a solution of 2-aminoacetonitrile and triethylamine in DMF and the mixture stirred at 25xc2x0 C. overnight. 1-Amino-cyclohexanecarboxylic acid cyanomethyl-amide may be recovered as described in the Examples. FMOC-1-aminocyclohexane carboxylic acid may be prepared as described in the Examples.
Compounds of the invention are either obtained in the free form, or as a salt thereof if salt forming groups are present.
Compounds of the Invention having basic groups can be converted into acid addition salts, especially pharmaceutically acceptable salts. These are formed, for example, with inorganic acids, such as mineral acids, for example sulfuric acid, a phosphoric or hydrohalic acid, or with organic carboxylic acids, such as (C1-C4)alkanecarboxylic acids which, for example, are unsubstituted or substituted by halogen, for example acetic acid, such as saturated or unsaturated dicarboxylic acids, for example oxalic, succinic, maleic or fumaric acid, such as hydroxycarboxylic acids, for example glycolic, lactic, malic, tartaric or citric acid, such as amino acids, for example aspartic or glutamic acid, or with organic sulfonic acids, such as (C1-C4)-alkylsulfonic acids (for example methanesulfonic acid) or arylsulfonic acids which are unsubstituted or substituted (for example by halogen). Preferred are salts formed with hydrochloric acid, methanesulfonic acid and maleic acid.
In view of the close relationship between the free compounds and the compounds in the form of their salts, whenever a compound is referred to in this context, a corresponding salt is also intended, provided such is possible or appropriate under the circumstances.
The compounds, including their salts, can also be obtained in the form of their hydrates, or include other solvents used for their crystallization.
The compounds of the invention exhibit valuable pharmacological properties in mammals and are particularly useful as inhibitors of cathepsin K.
The cathepsin K inhibitory effects of the compound of the invention can be demonstrated in vitro by measuring the inhibition of e.g. recombinant human cathepsin K.
The in vitro assay is carried out as follows:
For cathepsin K:
The assay is performed in 96 well microtiter plates at ambient temperature using recombinant human cathepsin K. Inhibition of cathepsin K is assayed at a constant enzyme (0.16 nM) and substrate concentration (54 mM Z-Phe-Arg-AMCA-Peptide Institute Inc. Osaka, Japan) in 100 mM sodium phosphate buffer, pH 7.0, containing 2 mM dithiothreitol, 20 mM Tween 80 and 1 mM EDTA. Cathepsin K is preincubated with the inhibitors for 30 min, and the reaction is initiated by the addition of substrate. After 30 min incubation the reaction is stopped by the addition of E-64 (2 mM), and fluorescence intensity is read on a multi-well plate reader at excitation and emission wavelengths of 360 and 460 nm, respectively. Compounds of the Invention typically have Kis for human cathepsin K of less than about 50 nM, preferably of about 5 nM or less, e.g. about 1 nM.
In view of their activity as inhibitors of cathepsin K, Compounds of the Invention are particularly useful in mammals as agents for treatment and prophylaxis of diseases and medical conditions involving elevated levels of cathepsin K. Such diseases include diseases involving infection by organisms such as pneumocystis carinii, trypsanoma cruzi, trypsanoma brucei, crithidia fusiculata, as well as parasitic diseases such as schistosomiasis and malaria, tumours (tumour invasion and tumour metastasis), and other diseases such as metachromatic leukodystrophy, muscular dystrophy, amytrophy and similar diseases.
Cathepsin K, has been implicated in diseases of excessive bone loss, and thus the Compounds of the Invention may be used for treatment and prophylaxis of such diseases, including osteoporosis, gingival diseases such as gingivitis and periodontitis, Paget""s disease, hypercalcemia of malignancy, e.g. tumour-induced hypercalcemia and metabolic bone disease. Also the Compounds of the Invention may be use for treatment or prophylaxis of diseases of excessive cartilage or matrix degradation, including osteoarthritis and rheumatoid arthritis as well as certain neoplastic diseases involving expression of high levels of proteolytic enzymes and matrix degradation.
Compounds of the Invention, are also indicated for preventing or treating coronary disease, atherosclerosis (including atherosclerotic plaque rupture and destabilization), autoimmune diseases, respiratory diseases and immunologically mediated diseases (including transplant rejection).
Compounds of the Invention are particularly indicated for preventing or treating osteoporosis of various genesis (e.g. juvenile, menopausal, post-menopausal, post-traumatic, caused by old age or by cortico-steroid therapy or inactivity).
Beneficial effects are evaluated in in vitro and in vivo pharmacological tests generally known in the art, and as illustrated herein.
The above cited properties are demonstrable in in vitro and in vivo tests, using advantageously mammals, e.g. rats, mice, dogs, rabbits, monkeys or isolated organs and tissues, as well as mammalian enzyme preparations, either natural or prepared by e.g. recombinant technology. Compounds of the Invention can be applied in vitro in the form of solutions, e.g. preferably aqueous solutions or suspensions, and in vivo either enterally or parenterally, advantageously orally, e.g. as a suspension or in aqueous solution, or as a solid capsule or tablet formulation. The dosage in vitro may range between about 10xe2x88x925 molar and 10xe2x88x929 molar concentrations. The dosage in vivo may range, depending on the route of administration, between about 0.1 and 100 mg/kg.
In accordance with the present invention it has been found that Compounds of the Invention, have good bioavailability, in particular good oral bioavailability. Thus, for example selected compounds of the Invention have absolute oral bioavailabilities of 50% or greater e.g. about 80% or more.
The antiarthritic efficacy of the Compounds of the Invention for the treatment of rheumatoid arthritis can be determined using models such as or similar to the rat model of adjuvant arthritis, as described previously (R. E. Esser, et. al. J. Rheumatology, 1993, 20, 1176.)
The efficacy of the compounds of the invention for the treatment of osteoarthritis can be determined using models such as or similar to the rabbit partial lateral meniscectomy model, as described previously (Colombo et al. Arth. Rheum. 1993 26, 875-886). The efficacy of the compounds in the model can be quantified using histological scoring methods, as described previously (O""Byrne et al. Inflamm Res 1995, 44, S117-S118).
The efficacy of the compounds of the invention for the treatment of osteoporosis can be determined using an animal model such as the ovariectomised rat or other similar species, e.g. rabbit or monkey, in which test compounds are administered to the animal and the presence of markers of bone resorption are measured in urine or serum (e.g. as described in Osteoporos Int (1997) 7:539-543).
Accordingly in further aspects the invention provides:
A Compound of the Invention for use as a pharmaceutical;
a pharmaceutical composition comprising a Compound of the Invention as an active ingredient;
a method of treating a patient suffering from or susceptible to a disease or medical condition in which cathepsin K is implicated, comprising administering an effective amount of a Compound of the Invention to the patient, and
the use of a Compound of the Invention for the preparation of a medicament for therapeutic or prophylactic treatment of a disease or medical condition in which cathepsin K is implicated.
The present invention relates to methods of using Compounds of the Invention and their pharmaceutically acceptable salts, or pharmaceutical compositions thereof, in mammals for inhibiting cathepsin K, and for the treatment of cathepsin K dependent conditions, such as the cathepsin K dependent conditions, described herein, e.g. inflammation, osteoporosis, rheumatoid arthritis and osteoarthritis.
Particularly the present invention relates to a method of selectively inhibiting cathepsin K activity in a mammal which comprises administering to a mammal in need thereof an effective cathepsin K inhibiting amount of a Compound of the Invention.
More specifically such relates to a method of treating osteoporosis, rheumatoid arthritis, osteoarthritis, and inflammation (and other diseases as identified above) in mammals comprises administering to a mammal in need thereof a correspondingly effective amount of a Compound of the Invention.
The following examples are intended to illustrate the invention and are not to be construed as being limitations thereon. Temperatures are given in degrees Centigrade. If not mentioned otherwise, all evaporations are performed under reduced pressure, preferably between about 15 and 100 mm Hg (=20-133 mbar). The structure of final products, intermediates and starting materials is confirmed by standard analytical methods, e.g. microanalysis and spectroscopic characteristics (e.g. MS, IR, NMR). Abbreviations used are those conventional in the art.